Our research proposal-Reductive Stress in the Pathogenesis of R120GCryAB Cardiomyopathy-is directly responsive to the new NHLBI Strategic Plan, which urges efforts to improve our understanding of the molecular and physiologic bases of heath and disease and the NIH Roadmap, which stipulates the creation of innovative collaborative research proposals between basic science and clinical scientists. The broad long-term objectives of this competitive renewal application are (i) to understand the pathogenic mechanisms by which hR120GCryAB expression-a genetic form of heart disease-promotes reductive stress and cardiomyopathy, and (ii) to develop therapeutic strategies targeted to similar molecular pathways for the prevention of heart failure in humans. Autosomal-dominant R120G in the gene encoding human 1B-crystallin (hR120GCryAB) causes a multisystem, protein aggregation disease including cardiomyopathy. Recent studies from our laboratory have shown that selective hR120GCryAB expression in the mouse heart induces such mechanism(s) linked to increased activity of glucose-6-phosphate dehydrogenase (G6PD) mimicking reductive stress. Reductive stress refers to an abnormal increase of reducing equivalents (e.g., glutathione, NADPH) but represents a continuum on the biological spectrum of redox homeostasis. This hypothesis-driven proposal will address four specific aims: namely, Aim 1. Determine if molecular interactions between mutant R120GCryAB and other proteins contribute to disease pathogenesis. We hypothesize that this might occur at sites of macromolecular complex formation involving cytosolic components, (e.g., Hsp25), a mutant chaperone and G6PD, providing a potential mechanism by which toxic gain-of function mutation leads to reductive stress;Aim 2. Determine the effects of hR120GCryAB expression on glutathione redox balance and cellular survival/death pathways. Aim 3. Determine if hR120GCryAB expression mediates cardiotoxicity by sustained activation of the redox-sensitive signaling pathways;and Aim 4: Characterize the impact of hR120GCryAB over-expression on mitochondrial function and myocardial substrate utilization. The following hypothesis is proposed: reductive stress, linked to hR120GCryAB over-expression, alters mitochondrial oxidative capacity and ATP synthesis, contributing to the development of heart failure. If successful, these efforts will improve our understanding about the molecular pathogenesis of genetic disorders, identify new therapeutic targets, and reduce the pain and suffering from cardiomyopathy and heart failure. PUBLIC HEALTH RELEVANCE: Autosomal dominant R120G in the gene encoding human 1B-crystallin causes cardiomyopathy (hR120GCryAB), a fatal condition characterized by enlargement (i.e., hypertrophy) of the heart and failure. Research from our laboratory has shown that transgenic mice mimicking the human condition were rescued when we engineered mice with much less glucose-6-phosphate dehydrogenase (G6PD). Our discovery establishes that G6PD plays a key role and supports the rationale for further studies that might lead to new therapies for human cardiac and degenerative diseases.